This invention relates to polysaccharides and bacterial nucleic acids which are capable of stimulating an immune response, and more particularly to substrate materials which potentiate that immune response. In specific and preferred embodiments of the invention, low molecular weight fragments of longer chain immune- stimulating polysaccharides and bacterial nucleic acids which have only a low bioactivity, as compared to the parent substance from which they are derived, are potentiated sufficiently so as to permit their use in substitution for the parent compound.
It is known that different uronic acid polymers with a .beta.1-4 glycosidic linkage are able to stimulate monocytes to produce tumour necrosis factor (TNF) through a membrane CD14 dependent manner (Espevik et al, Eur. J. Immunol. 23:255). Mannuronan (poly M) is the most potent of the .beta.1-4 linked uronic acid polymers in inducing cytokine production. However, the cytokine stimulatory activity of mannuronan is dependent of the molecular weight of the polymer, and optimal cytokine induction is obtained when the MW is 50,000 or higher (Otterlie et al, Infect. Immun. 61: 1993 pages 1917-1925. However, there is a sharp decline in activity at lower molecular weights and all useful activity is lost at a molecular weight below 10,000 g/mol. Although there are no apparent toxic effects when high molecular weight mannuronan is injected into mice, nonetheless it is important to use a polymer size as small as possible for therapeutic purposes in order to promote more complete and rapid excretion of the injected material from the body. This requirement therefore conflicts with the desire to optimise the TNF stimulating activity.
It is also known that lipopolysaccharide (LPS) has a TNF-inducing ability which depends on the three- dimensional supramolecular structure (Rietschel et al, "Bacterial endotoxins: properties and structure of biologically active domains", Werlag Chemie, 1988, p1). These supramolecular structures depend on the amount and distribution of the acyl chains in the lipid A region of LPS, and when lipid A occurs in a cubic or inverted hexagonal structure an increased cytokine induction is observed, whereas a lamella structure gives no cytokine induction. LPS as such is highly toxic but it can be delipidized by alkaline hydrolysis to form a detoxified LPS (D-LPS) from which the lipid A region, which is the main cause of the toxicity, has been removed. However, D-LPS has only a low ability to stimulate monocytes to produce TNF, despite retaining an intact polysaccharide portion.
It is disclosed by Seljelid et al of the Institute of Medical Biology, University of Troms.o slashed., Norway in Scand. J. Immunol. 25, 55-60, (1987) that plastic microbeads derivatized with .beta.-1,3-D glucan protect mice against pneumococcal and E. coli infections. These and other workers at the Institute of Medical Biology have subsequently shown that this protective effect is caused at least in part by stimulation of cytokine release (see, for example, Rasmussen et al, Journal of Cellular Biochemistry (1991) 46:60-68).
Other members of the family of polymers of uronic acid are also known to stimulate monocytes to produce TNF or other cytokines. For instance, D-glucuronic acid (D-GlcA) has this property, although with less potency compared with mannuronan.
Polysaccharides from gram-negative and gram- positive bacteria such as lipoarabinomannan, lipoteichoic acid and peptidoglycans are also known to induce cytokines.
Another well known immune-stimulating polysaccharide is chitosan.
It is, however, to be understood that not all polysaccharides have the ability to stimulate monocytes.
Certain bacterial nucleic acids form another category of substances with the capability of stimulating an immune response.